1. Field of the Invention
This invention relates to compositions and methods for detecting the presence of endogenous retroviruses, in particular, the detection of porcine endogenous retroviruses (PERV) in tissues useful as a xenograft.
2. General Background and State of the Art
The most common source of tissue used today as donor tissue for transplantation is the allograft (same species, different person). However, there are insufficient resources of human organs and cells for use as donor tissue. The shortage of human donor material has resulted in alternative solutions for transplantation. Xenotranplantation, which is the use of living tissue from non-human animals is one viable alternative. However, when using alternative sources, ethical, practical, biological and economic concerns must be considered. Therefore, among the animal species most suitable for use as a xenotranplant is the pig.
Examples of porcine tissue already being tested in clinical trials include fetal pig pancreatic islet cells for treating diabetes (Groth et al., Lancet 1994), pig neuronal cells for treating Parkinson's disease (Deacon et al., Nat Med 1997) and extracorporeal (ex vivo) pig liver or kidney perfusion for treating liver (Foley et al., Transplantation 2000, Levy et al., Transpl. 2000. 69: 272) or kidney failure (Breimer et al., Xenotransplantation 1996). However, as the potential for success increases the use of porcine tissue as a resource, the potential for introducing an infectious agent from the pig into the recipient becomes an increasing concern. Although risks associated with some pathogens can be reduced by breeding for and using specific-pathogen-free (SPF) animal colonies, this approach is not feasible for preventing infection from endogenous retroviruses, because these pathogens exist in the germine of all pigs.
Porcine Endogenous Retrovirus (PERV) is a C-type retrovirus that is permanently integrated in the pig genome. PERVs exist in the pig genome at an estimated 25–50 copies per cell. Early scientific reports dating back to the 1970's indicated that some porcine (pig) cells grown in culture produced type C retrovirus particles. This suggested their potential to be infectious. However, other studies of cells cultured directly from pig tissue (primary culture) showed no evidence of infectious potential. More recent reports indicate that there is a low frequency of PERV infection of human cells that are co-cultured with a pig cell line, PK15 (Le Tissier et al., Nature 1997; Patience et al., Nature Medicine, 1997). This reemphasized the concern regarding potential risk of PERV infection to human recipients of porcine tissue xenografts. Augmenting this risk is the use of immunosuppressive therapies necessary for preventing graft rejection by a recipient. Suppression of the recipient's immune system may also significantly increase the recipient's susceptibility to infection by PERV. Therefore, there is a need for methods to detect and monitor porcine tissues and cells for the presence of infectious PERV.
To date, there are three classes of PERV (PERV-A, PERV-B, and PERV-C). The differences between the three classes are primarily based on sequence differences in their envelope gene region. Recent identification of PERV sequences allows for development of molecular based detection methods. For example, methods that can detect specific sequences of DNA such as the polymerase chain reaction (PCR) can be used to identify the presence of PERV in a tissue sample. Keeping in mind that all pig genomes have PERV sequences, but that not all PERV are infectious, prescreening of a transplant tissue merely for presence of a PERV sequence is not a sufficient indicator of its infectious potential. Therefore, current methods for detecting the presence of PERV are limited by their inability to determine which PERV loci are infectious.
Thus, there is a need for methods and compositions capable of reducing the risk of transmission of PERV from porcine tissues suitable for use as xenografts. Particularly needed are compositions and methods for detecting the presence of infectious PERV in a biological sample.